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J Biol Chem. 2004 Jul 2;279(27):28113-21. Epub 2004 Apr 28.

Post-transcriptional regulation of sterol regulatory element-binding protein-1 by ethanol induces class I alcohol dehydrogenase in rat liver.

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1
Arkansas Children's Nutrition Center, University of Arkansas for Medical Sciences, 1120 Marshall Street, Little Rock, AR 72202, USA.

Abstract

Members of the sterol regulatory element-binding protein (SREBP) family of transcription factors control the synthesis and uptake of cholesterol, fatty acids, triglycerides, and phospholipids. Continuous intragastric infusion of ethanol-containing diets as part of total enteral nutrition generates well defined 6-day cycles (pulses) of urine ethanol concentrations (UECs) in rats. Pulsatile UECs are the result of cyclical expression and activity of the principal alcohol-metabolizing enzyme, hepatic Class I alcohol dehydrogenase (ADH), and this mechanism involves regulated CCAAT/enhancer-binding protein-beta expression and binding to the ADH promoter. In this study, we further explore the molecular mechanism for ethanol-induced ADH expression during the UEC pulse in adult male rats fed ethanol by total enteral nutrition for 21-30 days. In hypophysectomized rats, in which the ADH protein increased by approximately 6-fold, the nuclear form of SREBP-1 decreased by approximately 7-fold. Because the ADH promoter contains two canonical sterol response element (SRE) sites (-63 to -53 and -52 to -40 relative to the transcription start site), electrophoretic mobility shift assays were conducted using an ADH-specific SRE site. Hepatic nuclear protein binding decreased by 2.4-fold on the ascending limbs and by 3.6-fold on the descending limbs of UEC pulses (p < 0.05). The specificity of nuclear protein binding to the ADH-SRE site was confirmed using antibody and UV cross-link assays. The in vivo binding status of SREBP-1 to ADH-SRE sites, as measured by the chromatin immunoprecipitation assay, had a pattern very similar to the electrophoretic mobility shift assay results. Functional analysis of the ADH-SREs demonstrated these sites to be essential for ADH transcription. In vitro transcription assays demonstrated that depletion of the SREBP-1 protein from nuclear extracts increased transcription activity by approximately 5-fold and that the liver X receptor agonist T0901317 (a known activator of SREBP-1c transcription) reduced in vitro expression of ADH mRNA by 2-fold. We conclude that SREBP-1 is a negative regulator of the ADH gene and may work in concert with the CCAAT/enhancer-binding proteins to mediate ethanol induction of ADH in vivo.

PMID:
15123720
DOI:
10.1074/jbc.M400906200
[Indexed for MEDLINE]
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